Quantitative measurements of physiological ions at proximity of biological membranes are crucial methods to investigate metabolic processes and to identify unhealthy cells. In fact, the variation in concentration gradients is still predominant in medical studies—pH in cancer cells[1], calcium ions and neurotransmitters in neurons[2,3], or sodium and potassium in various excitable cells.[4-6] Current methods focus mainly on fluorescent markers to react to these environmental changes by a modification in their emissive properties in different conditions. However, these markers have often shown a certain cytotoxicity[7,8], which limits their use for long-term analyses and raises many questions concerning the stress induced during these tests.
Because of its intrinsic analytical advantages—namely, minimal photophysical stress induction, high sensitivity to minute signal variation, and adaptability on multiple biomedical platforms—fluorescence spectroscopy continues to be a dominant technology in various fields. Moreover, it has been found that dipole-dipole coupling with conductive electrons of a metallic surface can improve the optical properties of organic fluorophores.[9-11] This collective oscillation, termed “surface plasmon”, is induced by specific electromagnetic wavelengths and can be localized on nanometric conductive domains. Metal-enhanced fluorescence (MEF) is therefore influenced by the position of the molecule in the resulting electric field, and this distance dependency is well documented on metallic surfaces with a thin silica spacer.[9] In recent years, the development of MEF core-shell nanoparticles has been the subject of multiple studies and is now a whole theme in itself.[12-15] Easily dispersible in most solvents, various diagnosis applications have arisen from this type of highly-luminescent nanoparticular architecture.[16,17]
Although showing multiple advantages for homogeneous sensing in aqueous solutions, core-shell nanoparticles have also been shown to be functional on two-dimensional substrates to create fluorescent chip-based sensors with a higher surface ratio than continuous metallic films. The covalent grafting of nanoparticular sensors on transparent matrices, e.g. silica coverslip, is particularly valuable for bio-characterization using fluorescence microscopes. This methodology allows for multiple emitters on the same device without undesirable FRET and better control of the fluorophore-plasmonic core distance, whereas deposited metal surfaces are limited in these aspects. Furthermore, the development of planar devices allows for high-throughput and rapid analysis of liquids by microfluidic spectrofluorimetry[18], while also inhibiting the formation of plasmonic aggregates, which results in a highly homogeneous fluorescent biochip.
The grafting of proteins on the surface of metallic nanoparticles by click chemistry has been described in the literature.[19] The “click” method has also been used on lamellar silica substrates in order to add antibodies and polysaccharides for surface sensing.[20-23]
The present disclosure refers to a number of documents, the contents of which are herein incorporated by reference in their entirety.